Attachment 1 Specifications.docx

HMW DNA extraction from fish finclips and PacBio HiFi Sequencing Whole Genome Sequencing in Rainbow Trout

SPECIFICATION

Statement of Work:

The USDA ARS National Center for Cool and Cold-Water Aquaculture (NCCCWA) requires whole genome sequencing via Pacific Biosciences (PacBio) Sequel DNA sequencing systems of twelve (12) rainbow trout samples providing at least 8x genome coverage per sample (Genome size ~2.5Gb).

Contractor shall provide all services, supplies, equipment, and facilities necessary to extract high molecular weight (HMW) DNA from rainbow trout fin clips stored in 95% ethanol and provide sequencing of 12 genomic DNA samples in accordance with the required performance elements and benchmarks.

Schedule of Line Items:

The rainbow trout fin clip samples in 95% ethanol will be provided to the vendor by September 30, 2022.

REQUIREMENTS.

1. A minimum amount of 15 ug of rainbow trout HMW genomic DNA extracted by the vendor per fin tissue sample. DNA samples’ concentration will be estimated using Qubit;

2. PacBio DNA (whole genome) library preparation performed by the vendor following the best practices recommended by PacBio. DNA and library prep QC must include HiFi library size selection;

3. PacBio Sequel DNA sequencing systems to produce High Fidelity (HiFi) long reads using the circular consensus sequencing (CCS) mode;

4. Read Length: N50 is at least 15 Kb;

5. Data Output: At least 20 Gb of pass filter reads (>Q20) per sample. The output data files delivered in standard PacBio BAM files format;

6. Vendor must have in place a series of quality control (QC) steps during genomic DNA extractions, libraries preparation and genomic DNA sequencing. Information on library preparation and QC procedures must be documented and included in the final project report, including documentation of the actual fragment sizes used in libraries preps, number of SMRT cells, total output per cell and number of pass filter reads per cell;

7. Deliverables will include the DNA sequencing data for all samples, all raw sequence data files, and Final Project Report. Unused fin tissue samples and unused genomic DNA samples extracted by the vendor will be sent back to the government. All data must be provided via secure portal download OR in an external hard drive (including shipping and handling) to the USDA-ARS-NCCCWA.

Completion and return of all deliverables is required within 60 days from receipt of all fin clip samples from the government. The order number shall be included on all documentation and shipments.